Disruption and analysis of the clpB, clpC, and clpE genes in Lactococcus lactis: ClpE, a new Clp family in gram-positive bacteria

In the genome of the gram-positive bacterium Lactococcus lactis MG1363, we have identified three genes (clpC, clpE, and clpB) which encode Clp protein s containing two conserved ATP binding domains. The proteins encoded by two of the genes belong to the previously described ClpB and ClpC families. Th e clpE gene, however, encodes a member of a new Clp protein family that is characterized by a short N-terminal domain including a putative zinc bindin g domain (-CX2CX22CX2C-). Expression of the 83-kDa ClpE protein as well as of the two proteins encoded by clpB was strongly induced by heat shock and, while clpC mRNA synthesis was moderately induced by heat, we were unable t o identify the ClpC protein. When we analyzed mutants with disruptions in c lpB, clpC, or clpE, we found that although the genes are part of the L. lac tis heat shock stimulon, the mutants responded like wild-type cells to heat and salt treatments. However, when exposed to puromycin, a tRNA analogue t hat results in the synthesis of truncated, randomly folded proteins, clpE m utant cells formed smaller colonies than wild-type cells and clpB and clpC mutant cells. Thus, our data suggest that ClpE, along with ClpP, which rece ntly was shown to participate in the degradation of randomly folded protein s in L. lactis, could be necessary for degrading proteins generated by cert ain types of stress.